Paneth Cell-Specific TRPV6 Channel Plays a Crucial Role in Alcohol-Associated Dysbiosis of Gut Microbiota and Tissue Injury at the Gut-Liver Axis

Abstract

Introduction: Endotoxemia plays an essential role in alcohol-associated tissue injury. Two factors contributing to endotoxemia are gut microbiota dysbiosis and epithelial barrier dysfunction. Previous studies indicated that TRPV6 channel-mediated Ca2+ influx plays a crucial role in alcohol-induced epithelial tight junction (TJ) disruption, mucosal barrier dysfunction, and endotoxemia. Trpv6 knockout mice were resistant to alcohol-induced intestinal barrier dysfunction and modulated microbiota dysbiosis, raising the question of how TRPV6 regulates microbiota composition. In this study, we investigated the role of Paneth cell-specific TRPV6 in alcohol-induced microbiota dysbiosis and barrier dysfunction. Methods: Adult Trpv6fl/fl (WT) and Trpv6fl/fl-Defa6Cre (KO; Paneth cell-specific TRPV6 knockout) mice were fed a Lieber-DiCarli liquid diet with EtOH (0% 2d, 1% 2d, 2% 2d, 4% 1wk, 5% 1wk, & 6% 1wk). Control groups were pair-fed an isocaloric EtOH-free diet. Intestinal permeability was measured by vascular-to-luminal flux of FITC-inulin in vivo, and mucosal inflammation was assessed by RT-PCR for cytokines/chemokines. Plasma lipopolysaccharide (LPS) was measured to evaluate endotoxemia. Gut microbiota was analyzed by 16S rRNA sequencing of colonic flushing. Lipidome in colonic flushing was analyzed using nontargeted liquid chromatography-mass spectrometry. Microbiome and lipidome data were processed by bioinformatic analyses. Results and conclusion: Data show that colon length was reduced, and colonic mucosal permeability in vivo was elevated by EtOH in WT mice. The increase in mucosal permeability was associated with increased plasma LPS levels. EtOH feeding elevated the expression of IL-6, IL-1β, and TNFα in the colon. EtOH-induced changes in colon length, mucosal permeability, cytokine expression, and plasma LPS were absent in KO mice. 16S rRNA sequencing data indicated that the microbiota composition in KO mice differed from that of WT mice. The relative abundance of Prevotellaceae, Xanthomonadaceae, and Streptococaceae were higher, and Psuedomonadaceae and Verrucomicrobiaceae abundance were lower in KO mice compared to WT mice. EtOH-induced increase in the abundance of Norcardiaceae and Psuedomonadaceae was low or absent in KO mice. EtOH increased Rhodococcus, Odoribacter, and Pseudomonas abundance at the genus level in WT but not KO mice. On the other hand, Clostridium, Turicibacter, and Helicobacter abundance were increased by EtOH in KO but not WT mice. Lipidomic analysis of colonic flushing showed that phospholipid contents were higher in KO mice than WT mice, irrespective of sex and treatments. A cluster of sphingolipids was dramatically high in KO female mice, which was reduced by EtOH feeding. Short-chain fatty acid esters of hydroxy fatty acids (AAHFA 3:0/24:0;O) were elevated by EtOH in male WT mice but not in female WT and male or female KO mice. In conclusion, these data indicate that Paneth cell-specific deletion of TRPV6 attenuates alcohol-induced changes in gut microbiota, colonic luminal lipids, mucosal barrier function, and endotoxemia in mice. NIH/NIAAA, RO1-AA029270; Veterans Administration IO1-BX003014. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.